Hostname: page-component-848d4c4894-8kt4b Total loading time: 0 Render date: 2024-06-19T09:11:08.294Z Has data issue: false hasContentIssue false
  • English
  • Français

Caprine blastocyst formation following intracytoplasmic sperm injection and defined culture

Published online by Cambridge University Press:  26 September 2008

L. Keskintepe ,
P.C. Morton ,
S.E. Smith ,
M.J. Tucker ,
A.A. Simplicio  and
B.G. Brackett
L. Keskintepe
Affiliation:
College of Veterinary Medicine, Athens, Georgia, USA, and Reproductive Biology Associates, Atlanta, Georgia, USA
P.C. Morton
Affiliation:
College of Veterinary Medicine, Athens, Georgia, USA, and Reproductive Biology Associates, Atlanta, Georgia, USA
S.E. Smith
Affiliation:
College of Veterinary Medicine, Athens, Georgia, USA, and Reproductive Biology Associates, Atlanta, Georgia, USA
M.J. Tucker
Affiliation:
College of Veterinary Medicine, Athens, Georgia, USA, and Reproductive Biology Associates, Atlanta, Georgia, USA
A.A. Simplicio
Affiliation:
College of Veterinary Medicine, Athens, Georgia, USA, and Reproductive Biology Associates, Atlanta, Georgia, USA
B.G. Brackett*
Affiliation:
College of Veterinary Medicine, Athens, Georgia, USA, and Reproductive Biology Associates, Atlanta, Georgia, USA
*
Dr Benjamin G. Brackett, The University of Georgia, College of Veterinary Medicine, Department of Physiology and Pharmacology, Athens, GA 30602, USA. Telephone: +1 (706) 542-5859. Fax: +1 (706) 542-3015. e-mail: brackett@calc.vet.uga.edu.
Get access Rights & Permissions [Opens in a new window]

Summary

Experiments were undertaken to develop intracytoplasmic sperm injection (ICSI) to produce caprine embryos out of the normal breeding season. Oocytes were obtained from 2–6 mm ovarian follicles at slaughter. Selected oocytes with two to four layers of cumulus cells were incubated in 1 ml of H-TCM 199 supplemented with 10 μg each of oFSH and bLH (NHPP, NIDDK, NICHD, USDA) and 20% fetal bovine serum (FBS) in a thermos (38.5°C) for 4.5 h during transportation. Then, oocytes were transferred into 75 μl of freshly prepared maturation medium under paraffin oil and a mixture of 5% O2, 5% CO2 and 90% N2. Approximately 26 h after recovery oocytes were denuded by incubation with hyaluronidase (100 IU/ml) and pipetting and held at 38.5°C for 90 min. Spermatozoa frozen in egg yolk extender were thawed in a 37°C water bath for 15s. Motile fractions were selected by swim-up, then incubated for 90 mm in TALP with 10 μg heparin/ml. Each oocyte was positioned with its first polar body at 6 or 12 o'clock by a holding pipette. Sperm (1 μl) were added to 10 μl medium containing 10% polyvinylpyrrolidone. A sperm cell was aspirated into a pipette, and then injected head-first into the cytoplasm of an oocyte maintained in H-TCM 199 + 20% FBS at 37°C. Injected oocytes were transferred to HM and, after 90 min, cultured in 50 μl of BSA-free synthetic oviduct fluid plus polyvinyl alcohol, citrate and non-essential amino acids. Results demonstrate that caprine blastocysts can be produced outside the breeding season by the use of frozen-thawed semen and injection of sperm cells with broken tails into ova followed by culture in defined medium.

Keywords

BlastocystCaprineDefined conditionsICSI
Type
Article
Information
Zygote , Volume 5 , Issue 3 , August 1997 , pp. 261 - 265
Copyright
Copyright © Cambridge University Press 1997

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Ebert, K.M., Selgarth, J.P., DiTullio, P., Denman, J., Smity, T.H., Memon, J.P., Schindler, J.E., Monastersky, G.M., Vitale, J.A. & Gordon, K. (1991). Transgenic production of a variant of human tissue-type plasminogen activator in goat milk: generation of transgenic goats and analysis of expression. Biotechnology 9, 835–8.Google ScholarPubMed
Goto, K., Konishita, A., Takuma, Y. & Ogawa, K. (1990). Fertilization in sperm injection in cattle (abstract). Theriogenology 33, 238.CrossRefGoogle Scholar
Iritani, A. (1990). Micromanipulation of oocytes and embryos. In Proceedings of the XIIIth World Congress on Fertility and Sterility,Marakesh, Morocco,1–5 October 1989, pp. 145–9. England:Parthenon Publishing.Google Scholar
Iritani, A. (1991). Micromanipulation of gametes for in vitro assisted fertilisation. Mol. Reprod. Dev. 28, 199–207.CrossRefGoogle Scholar
Iritani, A., Utsumi, K., Miyake, M., Hosoi, Y. & Saeki, K. (1988). In vitro fertilisation by a routine method and micromanipulation. In In Vitro Fertilisation and Other Assisted Reproduction, ed. Jones, H. & Shrader, C.. Ann. N.Y. Acad. Sci. 541, 583–90.CrossRefGoogle Scholar
Jannny, L. & Menezo, Y.J.R. (1994). Evidence for a strong paternal effect on human preimplantation embryo development and blastocyst formation. Mol. Reprod. Dev. 38, 36–42.CrossRefGoogle Scholar
Keefer, C.L. (1989). Fertilisation by sperm injection in the rabbit. Gamete Res. 22, 59–69.CrossRefGoogle ScholarPubMed
Keefer, C.L., Younis, A.I. & Brackett, B.G. (1990). Cleavage development of bovine oocytes fertilized by sperm injection. Mol. Reprod. Dev. 25, 265–76.CrossRefGoogle ScholarPubMed
Keskintepe, L. & Brackett, B.G. (1996). In vitro developmental competence of in vitro matured bovine oocytes fertilized and cultured in completely defined media. Biol. Reprod. 55, 333–9.CrossRefGoogle ScholarPubMed
Keskintepe, L., Burnley, C.A. & Brackett, B.G. (1995). Production of viable bovine blastocysts in defined in vitro conditions. Biol. Reprod. 52, 1410–17.CrossRefGoogle ScholarPubMed
Keskintepe, L., Darwish, G.M., Kenimer, A.T. & Brackett, B.G. (1994). Term development of caprine embryos derived from immature oocytes in vitro. Theriogenology 42, 527–35.CrossRefGoogle ScholarPubMed
Keskintepe, L., Luvoni, G.C., Rzucidlo, S.J. & Brackett, B.G. (1996). Procedural improvements for in vitro production of viable uterine stage caprine embryos. Small Ruminant Res. 20, 247–54.CrossRefGoogle Scholar
Markert, C.L. & Peters, R.M. (1977). Homozygous mouse embryos produced by microsurgery. J. Exp. Zool. 228, 195–201.CrossRefGoogle Scholar
Martino, A., Mogas, T., Palomo, M.J. & Paramio, M.T. (1995). In vitro maturation and fertilization of prepubertal goat oocytes. Theriogenology 43, 473–85.CrossRefGoogle ScholarPubMed
Palermo, G., Joris, H., Devroey, P. & Van Steirteghem, A. (1992). Pregnancies after intracytoplasmic sperm injection of single spermatozoon into an oocyte. Lancet 340, 17–18.CrossRefGoogle ScholarPubMed
Simplicio, A.A., Brackett, B.G. & Keskintepe, L. (1997). Use of cryopreserved spermatozoa for caprine in vitro fertilization (UVF) (abstract). Theriogenology 47, 299.CrossRefGoogle Scholar
Trounson, A. & Sathananthan, A.H. (1993). Fertilisation using micromanipulation techniques. In Handbook of In Vitro Fertilisation, ed. Trounson, A. & Gardner, D.K., pp. 131–50. Boca Raton, FL: CRC Press.Google Scholar
Tucker, M.J., Wright, G., Morton, P.C., Mayer, M.P., Ingargiola, P.E. & Jones, A.E. (1995). Practical evolution and application of direct intracytoplasmic sperm injection for male factor and idiopathic fertilisation failure infertilities. Fertil. Steril. 63, 820–7.CrossRefGoogle ScholarPubMed
Uehara, T. & Yanagimachi, R. (1977). Behavior of nuclei of testicular, caput and cauda epididymal spermatozoa injected into hamster eggs. Biol. Reprod. 16, 315–21.CrossRefGoogle ScholarPubMed
Younis, A.I., Keefer, C.L. & Brackett, B.G. (1989). Fertilisation of bovine oocytes by sperm injection (abstract). Theriogenology 31, 276.CrossRefGoogle Scholar